Voltage-gated ion channels (VGICs) are involved in electrical signal transduction and play essential roles in life activities. How membrane potential drives conformational change at the voltage-sensing domain (S1-S4, VSD) and regulates channel gating is a central theme for voltage-gated ion channels. To elucidate voltage-gating mechanisms, one needs to capture VSD structures in both the activated state and resting state.

In recent years, the research team led by Prof. GUO Jiangtao from Zhejiang University School of Medicine has conducted systematic research into the voltage gating mechanism of TPC1. The latest research results were published in the journal PNAS on November 30, and entitled “Voltage-gating and cytosolic Ca²⁺ activation mechanisms of Arabidopsis two-pore channel AtTPC1”.

AtTPC1 belongs to the VGIC superfamily. It is localized in the vacuolar membrane and is responsible for generating slow vacuolar (SV) current. In addition to being activated by membrane depolarization, AtTPC1 is also doubly regulated by Ca²⁺. It is activated by cytosolic Ca²⁺ binding at the EF-hand domain but inhibited by vacuolar Ca²⁺ binding at VSDII. In order to obtain the structure of AtTPC1 in open-state with the presence of Ca²⁺, based on the previous study, researchers used the mutant AtTPC1ΔCai with three Ca²⁺-coordinating residues on VSDII (D240A/D454A/E528A), which can mitigate the vacuolar Ca²⁺ inhibition.